Francisella tularensis is a pathogenic intracellular bacterium capable of causing infectious disease in more than 150 mammalian species. Arthropod vectors, such as ticks, flies and mosquitoes, are frequently involved in the transmission of the pathogen to mammals but it can be transmitted also via contaminated food and water, and aerosols. There are four subspecies of F. tularensis, but only two, subspecies tularensis (type A), and subspecies holarctica (type B), are commonly infectious for humans. Only type A strains of F. tularensis may cause lethal infection in humans, in particular untreated respiratory tularemia has a high mortality rate if left untreated.
Because of its high infectivity, ease of dissemination by aerosol, and capacity to cause severe morbidity and mortality, type A F. tularensis has long been considered a potential biological warfare agent. However, to date no specific virulence factors that explain the high virulence of type A strains have been identified. A comparative genomic analysis showed that the proportion of genes conserved among the four subspecies of F. tularensis is high, >97%, and that less than 30 of a total of 1,800 genes are unique to type A versus type B F. tularensis. 
Live attenuated F. tularensis vaccines were developed in Russia in the 1950's from a type B strain. One of these strains, designated as the live vaccine strain, LVS, was transferred to the US. Vaccine studies conducted on volunteers in the 1960's demonstrated that it protected humans against systemic inoculation or inhalation of a type A strain of the pathogen. Epidemiological studies of tularemia cases among Francisella researchers before and after the introduction of LVS vaccination confirmed its utility. However, despite having been developed almost 50 years ago, the nature of the genetic lesion responsible for its attenuation, the protective antigens, and the immunological basis for its efficacy remain unknown.
Moreover, in both human and animal studies, systemic vaccination with LVS provided sub-optimal protection against aerosol challenge with type A F. tularensis. For these reasons, LVS has never been licensed as a vaccine. In the past, it was granted investigational new drug (IND) status, but this was revoked by the FDA several years ago. These problems with LVS have motivated a search for better-defined vaccines of equal or greater efficacy.
Because the protective protein antigens of F. tularensis are completely unknown, a sub-unit vaccine is currently inconceivable, especially as it would need to be formulated with an adjuvant system able to elicit robust CD4+ and CD8+ T cell responses, both of which are known to be required to control tularemia. Although experimental adjuvants with these properties exist, none have yet been approved for clinical use.
The identities and characteristics of the virulence factors and protective antigens of the subspecies of Francisella are essentially unknown. Although recent comparative genomics analyses have begun to demonstrate genetic differences among the subspecies, these alone have been insufficient to explain their relative virulence. Moreover, it is known that sublethal infection of mice with subspecies novicida fails to confer protection against subsequent challenge with subspecies holarctica or tularensis suggesting that the protective antigens are highly restricted. It has also been observed that only certain mouse strains can be protected by LVS. On this basis, it seems unlikely that defined mutants of the holarctica subspecies would be more effective vaccines than LVS.
Thus, it would be desirable to generate a defined type A strain mutant lacking the minimum number of genes required to render it a safe and effective live vaccine. However, no successful strategy has been disclosed in the prior art.